Project summary/abstract The alternating contraction of opposing flexor and extensor muscles, known as antagonist pairs, creates the rhythmic limb movement of locomotion. This phenomenon is regulated in part by reciprocal inhibition: sensory feedback from an active muscle excites the Ia interneuron, which then inhibits that muscle's antagonist. However, when a task requires limb stiffness and joint stability, this circuit must be overridden to allow co-contraction of both flexor and extensor muscles. Previous studies have indicated that motor cortex is responsible for the reduction of reciprocal inhibition observed during voluntary co-contraction, but its mechanism of action is unknown. Elucidating how motor cortex recruits spinal circuits to permit antagonist muscle co-contraction will further our understanding of the neural control of voluntary movement. Monkey and cat studies have reported that intracortical inhibition is reduced during voluntary co- contraction, indicating that this reduction may be necessary for co-contraction. Neural recording studies in monkey found that a discrete population of corticospinal neurons (CSNs) increases its activity during co- contraction but not during extension or flexion, indicating that increased CSN activity may be required for this behavior. Of the CSNs, a subgroup that synapses on a type of spinal interneuron known as the GABApre (CSN-GABApres) is a likely candidate for antagonist muscle control. This is supported by findings that indicate the GABApre interneuron is capable of reducing reciprocal inhibition and that the type of inhibition GABApres exert is increased during co-contraction. This evidence leads us to hypothesize that during this behavior, intracortical inhibition is decreased, the activity of CSNs, in particular CSN-GABApres, is increased, and that this activity is necessary for voluntary co-contraction. To test these hypotheses, we will record motor cortical activity in mouse during a novel behavioral paradigm that elicits either co-contraction or alternation of the forelimb triceps-biceps antagonist muscle pair. Putative cortical interneurons will be identified by the width of their action potential waveform and CSNs will be identified by optogenetic activation of their axons. A novel tracing technique will also allow the optical identification of CSN-GABApres during recording. The importance of CSN and CSN-GABApre activity to the reduction of spinal reciprocal inhibition and thus the execution of co-contraction will be tested by optogenetic inactivation of these cells during co-contraction as compared to alternation. The findings generated by these experiments will clarify the neural mechanisms that underlie the control of antagonist muscles and voluntary movement. This information could eventually be applied to treatment for stroke and spinal cord injury patients or contribute to the development of neural prostheses for movement-impaired individuals.